Substantially continuous attention is focused on the development of improved contraceptive methods. One widely exploited technology is the use of spermicides, essentially a chemical barrier that prevents penetration of sperm to the uterus or egg, or inhibits the activity thereof, thereby precluding fertilization One of the most widely used spermicides is a detergent, Nonoxynol-9. Reports indicate an increased incidence of urogenital infections and cervicovaginal inflammation in women employing this detergent spermicide. McGroarty et al, Journal of Urology, 152(3):831-833 (1994).
As an alternative to chemical detergents, authors have suggested the use of monoclonal antibodies as likely safe active agents for topical applications, such as use in topical spermicides. See, e.g., Cone et al, Am. J. Reprod. Immunol., 32:114-131 (1994). Studies conclude that in addition to the reduction or elimination of unwanted immune reactions, human monoclonal antibodies should present safe spermicides since their dose and duration of application can be readily controlled, topical delivery minimizes systemic exposure and the monoclonal antibody can be selected for safety and efficacy. Therefore, a sperm-active monoclonal antibody delivered as a topical spermicide may produce desired anti-fertility effects without the negative side effects accompanied by detergent spermicides. See generally, Alexander, Scientific American, September:136-141 (1995). Accordingly, a goal in the art continues to be the provision of a safe and effective spermicide employing monoclonal antibodies.
Many investigators around the world are looking at the possibility of the development of contraceptive vaccines based on sperm antigens. See, e.g., Aitken et al, British Medical Journal, 49:88-99 (1993), Preemerman et al, Biol. Reprod., 50:615-621 (1994) and Herr, Fertility Control, pp. 431-452 (Second Edition 1994). In this connection, work continues on human chorionic gonadotropin (hCG) as a contraceptive vaccine for women. Talwar, Current Opinion in Immunology, 6:698-704 (1994) and European Patent 86304274.3. While clinical vaccine trials are underway with this potential vaccine, the hCG immunogen employed functions as an abortifactant, that is, immune responses induced by inoculation with this vaccine induce abortion of the early embryo or fetus. This may constitute an unacceptable form of contraceptive for many individuals.
As an alternative, a variety of sperm surface antigens have been employed in studies involving primate and rodent models. Thus, decreased fertility rates resulted from the immunization of test animals with sperm surface antigens such as LDH-C4, O'Hern et al, Biol. Reprod., 52:331-339 (1995), PH-20, Primakoff et al, Nature, 335:543,546 (1988), RSA-1, O'Rand et al, J. Reprod. Immunol., 25:89-102 (1993) and fertilin, Ramarao et al, Mol. Reprod. Dev., 43:70-75 (1995). Disappointingly, in primates, the highest rate of efficacy observed with a sperm antigen is about 75 percent inhibition of fertility, O'Hern et al, supra. Thus to date there has not been identified a human sperm antigen that functions as a contraceptive vaccine with a level of efficacy comparable to that of oral contraception. Thus, it remains an object of those of skill in the art to provide a safe and effective contraceptive vaccine with a high rate of fertility inhibition, on the order of the level of efficacy given by oral contraceptives.
Additionally, because those receiving a contraceptive vaccine will require periodic monitoring of serum antibody to determine if they are “safe”, use of the sperm specific antigen as a target in assays to measure antibody concentration in persons receiving the vaccine is desirable.
“Over the counter” assay or diagnostic kits for the detection of hormones associated with pregnancy (hCG and others) have achieved wide-spread success in the marketplace, as an alternative or a first-step to potentially embarrassing, inconvenient and expensive visits to medical offices. In recent years, attention has been focused on assays for the presence, and concentration of sperm in a users ejaculate. Both from the point of view of fertility counseling, as well as clinical diagnosis in the case of rape, or for the purposes of assaying for the presence and effectiveness of a vasectomy, a convenient test kit, that could be safely and reliably employed at home, for the detection of sperm in a sample, has become increasingly desirable. In accordance with the present invention, a test kit employing a monoclonal antibody against any of the sperm antigens disclosed herein, is also within the scope of the present invention.
Purification of the sperm specific antigens is the first step in preparation of an effective vaccine. The purified antigen, incorporated in a pharmaceutically acceptable carrier, can be administered to patients desiring vaccination for contraception. Repeated vaccination results in the generation of antibodies against sperm, highly effective in the binding of sperm. To monitor the development of an effective level of antibodies, the purified antigen may be used as a test standard reagent, to determine the presence and amount of antibodies present in the patient, through conventional diagnostics.
Definitions
In describing and claiming the invention, the following terminology will be used in accordance with the definitions set forth below.
As used herein, “nucleic acid,” “DNA,” and similar terms also include nucleic acid analogs, i.e. analogs having other than a phosphodiester backbone. For example, the so-called “peptide nucleic acids,” which are known in the art and have peptide bonds instead of phosphodiester bonds in the backbone, are considered within the scope of the present invention.
The term “peptide” encompasses a sequence of 3 or more amino acids wherein the amino acids are naturally occurring or synthetic (non-naturally occurring) amino acids. Peptide mimetics include peptides having one or more of the following modifications:                1. peptides wherein one or more of the peptidyl —C(O)NR— linkages (bonds) have been replaced by a non-peptidyl linkage such as a —CH2-carbamate linkage (—CH2OC(O)NR—), a phosphonate linkage, a —CH2-sulfonamide (—CH2—S(O)2NR—) linkage, a urea (—NHC(O)NH—) linkage, a —CH2-secondary amine linkage, or with an alkylated peptidyl linkage (—C(O)NR—) wherein R is C1-C4 alkyl;        2. peptides wherein the N-terminus is derivatized to a —NRR1 group, to a —NRC(O)R group, to a —NRC(O)OR group, to a —NRS(O)2R group, to a —NHC(O)NHR group where R and R1 are hydrogen or C1-C4 alkyl with the proviso that R and R1 are not both hydrogen;        3. peptides wherein the C terminus is derivatized to —C(O)R2 where R 2 is selected from the group consisting of C1-C4 alkoxy, and —NR3R4 where R3 and R4 are independently selected from the group consisting of hydrogen and C1-C4 alkyl.        
Naturally occurring amino acid residues in peptides are abbreviated as recommended by the IUPAC-IUB Biochemical Nomenclature Commission as follows: Phenylalanine is Phe or F; Leucine is Leu or L; Isoleucine is Ile or I; Methionine is Met or M; Norleucine is Nle; Valine is Vat or V; Serine is Ser or S; Proline is Pro or P; Threonine is Thr or T; Alanine is Ala or A; Tyrosine is Tyr or Y; Histidine is H is or H; Glutamine is Gln or Q; Asparagine is Asn or N; Lysine is Lys or K; Aspartic Acid is Asp or D; Glutamic Acid is Glu or E; Cysteine is Cys or C; Tryptophan is Trp or W; Arginine is Arg or R; Glycine is Gly or G, and X is any amino acid. Other naturally occurring amino acids include, by way of example, 4-hydroxyproline, 5-hydroxylysine, and the like.
Synthetic or non-naturally occurring amino acids refer to amino acids which do not naturally occur in vivo but which, nevertheless, can be incorporated into the peptide structures described herein. The resulting “synthetic peptide” contain amino acids other than the 20 naturally occurring, genetically encoded amino acids at one, two, or more positions of the peptides. For instance, naphthylalanine can be substituted for trytophan to facilitate synthesis. Other synthetic amino acids that can be substituted into peptides include L-hydroxypropyl, L-3,4-dihydroxyphenylalanyl, alpha-amino acids such as L-alpha-hydroxylysyl and D-alpha-methylalanyl, L-alpha.-methylalanyl, beta.-amino acids, and isoquinolyl. D amino acids and non-naturally occurring synthetic amino acids can also be incorporated into the peptides. Other derivatives include replacement of the naturally occurring side chains of the 20 genetically encoded amino acids (or any L or D amino acid) with other side chains.
As used herein, the term “conservative amino acid substitution” are defined herein as exchanges within one of the following five groups:                I. Small aliphatic, nonpolar or slightly polar residues:                    Ala, Ser, Thr, Pro, Gly;                        II. Polar, negatively charged residues and their amides:                    Asp, Asn, Glu, Gln;                        III. Polar, positively charged residues:                    His, Arg, Lys;                        IV. Large, aliphatic, nonpolar residues:                    Met Leu, Ile, Val, Cys                        V. Large, aromatic residues:                    Phe, Tyr, Trp                        
As used herein, the term “purified” and like terms relate to the isolation of a molecule or compound in a form that is substantially free of contaminants normally associated with the molecule or compound in a native or natural environment.
As used herein, the term “C7/8 polypeptide” and like terms refers to polypeptides comprising SEQ ID NO: 2 and biologically active fragments thereof.
As used herein, the term “SAMP32 polypeptide” and like terms refers to polypeptides comprising SEQ ID NO: 9 and biologically active fragments thereof.
As used herein, the term “C58 polypeptide” and like terms refers to polypeptides comprising SEQ ID NO: 16 and biologically active fragments thereof.
As used herein, the term “biologically active fragments” or “bioactive fragment” of an C7/8, SAMP32 or C58 polypeptide encompasses natural or synthetic portions of those polypeptides that are capable of specific binding to at least one of the natural ligands of the native polypeptide.
“Operably linked” refers to a juxtaposition wherein the components are configured so as to perform their usual function. Thus, control sequences or promoters operably linked to a coding sequence are capable of effecting the expression of the coding sequence.
As used herein, the term “pharmaceutically acceptable carrier” encompasses any of the standard pharmaceutical carriers, such as a phosphate buffered saline solution, water and emulsions such as an oil/water or water/oil emulsion, and various types of wetting agents.